Camier S, Dechampesme A M, Sentenac A
Service de Biochimie et de Génétique Moléculaire, Commissariat à l'Energie Atomique, Saclay, Gif-sur-Yvette, France.
Proc Natl Acad Sci U S A. 1995 Sep 26;92(20):9338-42. doi: 10.1073/pnas.92.20.9338.
We have developed a system to transcribe the yeast 5S rRNA gene in the absence of the transcription factor TFIIIA. A long transcript was synthesized both in vitro and in vivo from a hybrid gene in which the tRNA-like promoter sequence of the RPR1 gene was fused to the yeast 5S RNA gene. No internal initiation directed by the endogenous 5S rDNA promoter or any processing of the hybrid transcript was observed in vitro. Yeast cells devoid of transcription factor TFIIIA, which, therefore, could not synthesize any 5S rRNA from the endogenous chromosomal copies of 5S rDNA, could survive if they carried the hybrid RPR1-5S construct on a multicopy plasmid. In this case, the only source of 5S rRNA was the precursor RPR1-5S transcript that gave rise to two RNA species slightly larger than wild-type 5S rRNA. This establishes that the only essential function of TFIIIA is to promote the synthesis of 5S rRNA. However, cells devoid of TFIIIA and surviving with these two RNAs grew more slowly at 30 degrees C compared with wild-type cells and were thermosensitive at 37 degrees C.
我们开发了一种在没有转录因子TFIIIA的情况下转录酵母5S rRNA基因的系统。从一个杂交基因在体外和体内均合成了一个长转录本，该杂交基因中RPR1基因的tRNA样启动子序列与酵母5S RNA基因融合。在体外未观察到由内源性5S rDNA启动子指导的内部起始或杂交转录本的任何加工过程。缺乏转录因子TFIIIA的酵母细胞因此无法从5S rDNA的内源性染色体拷贝合成任何5S rRNA，但如果它们在多拷贝质粒上携带杂交RPR1-5S构建体，则能够存活。在这种情况下，5S rRNA的唯一来源是前体RPR1-5S转录本，它产生了两种比野生型5S rRNA略大的RNA物种。这表明TFIIIA的唯一基本功能是促进5S rRNA的合成。然而，缺乏TFIIIA并靠这两种RNA存活的细胞在30℃下比野生型细胞生长得更慢，并且在37℃下对温度敏感。
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